Test series with living cells are of paramount importance in biological and medical research and development, and are used to a large extent in the pharmaceutical industry, for example in the development of new active substances and medicaments. In this context there is a requirement for observing a multitude of different cell probes quickly and as far as possible in an automated manner by means of transmission microscopy or fluorescence microscopy, and during observation or between subsequent observation steps to feed, in a controlled manner, to the individual probes liquid substances which can, for example, comprise an active medical substance or some molecular-biological modification (e.g. siRNA or immunostaining).
To this purpose cell probes are distributed to separate chambers of an object carrier or of a multiwell plate, and for observation said probes, on a stage, are sequentially fed into the optical path of a microscope. In this process inverse microscopes are frequently used in which the image recording device and often also at least part of the illumination device are arranged below the stage so that the space above the stage can predominantly be reserved for positioning and filling the probe. For quick automatic positioning of the probes, generally speaking probe positioners are provided on the stage, which probe positioners can accommodate a multitude of different probe containers, wherein the individual probe chambers of said probe positioners can quickly and very accurately be moved into the optical path of the microscope, and for focussing can also be displaced along the optical axis. Adding a liquid, for example an active substance, at a determined dilution ratio or mixing ratio to the individual probe chambers, or removing the liquid by suction from the probe chambers, in part still takes place manually, for example with the use of a microliter pipette. However, manual filling or removal by suction is not only time-consuming and labour-intensive, but also quite error-prone.
Furthermore, it is often necessary to carry out test series under controlled temperature conditions and atmospheric conditions so that the probe container, together with the stage and parts of the observation equipment and illumination device, is accommodated in a so-called climatic chamber, and consequently the probes are practically no longer accessible from the exterior. For this reason automated pipetting systems with movable pipettes were invented, which systems make it possible to quickly and reliably feed a liquid into selected pipetting containers in a controlled atmosphere. In this arrangement, the liquid infeed from a reservoir normally takes place with the use of pumps comprising micro motors; positioning and operation of the pipette also takes place with the use of electric motors. An inverse microscope with such an automated pipetting device within a climatic chamber is, for example, described in patent specification U.S. Pat. No. 7,092,151 B2.
The pipetting devices known from prior art are associated with a difficulty in that the pipette and the drive unit require comparatively large installation space, and as a result of this they could negatively affect operation of the microscope. This problem occurs in particular if the probes in transmission are to be illuminated from above the stage, because the illumination device and the pipetting device cause mutual interference. However, the same difficulties can also arise in test series in which the illumination can be exclusively from below the stage, while the microscope used normally provides for a second illumination device above the stage, which second illumination device, while not needed for the test series that is to be carried out, nonetheless for design reasons cannot be moved out of the optical path far enough to allow unrestricted operation of the automatic pipetting device. Many widely used microscope types are associated with such spatial restrictions and can therefore be used with automatic pipetting devices known from prior art only with functional limitations or only after possibly time-consuming and cost-intensive modifications. There is thus a requirement for a pipetting device that can be used without mutual interference with microscopes of a known and widely used design.
Furthermore, the probe chambers used are often sealed by means of a cover, for example a metal foil or a plastic film, in order to protect the probe from the ambient atmosphere. There is thus furthermore a requirement for an automatic pipetting device which despite such a sealing arrangement makes it possible to quickly, reliably and precisely feed a liquid into selected probe chambers.
Likewise, apart from allowing automated feed-in of a liquid into selected probe chambers, the pipetting device should also allow the quick and effective removal of a liquid from selected probe chambers.
These objects are met by the coaxial needle according to the invention according to claim 1, or by the pipetting device according to the invention according to claim 5 and claim 11. The invention also relates to the corresponding pipetting method according to claim 13. The subordinate claims relate to preferred embodiments.